The present invention relates to a concentrate of thrombin in coagulable proteins, a method of obtaining the same from total plasma and a use thereof for therapeutic purposes.
This type of concentrate may be used for obtaining, by redissolution, an injectable fibrinogen or a biological glue.
As is known, injections of fibrinogen make it possible to treat hypofibrinogenesis or constitutional afibrinogenesis conditions, with or without hemorrhages, and also the acute defibrination syndromes with serious hemorrhages, in association with etiological treatment, heparinotherapy or antifibrinolyties.
On the other hand, biological glues make it possible to give efficient help during certain clinical episodes such as skin grafts, nerve or artery sutures, rapid cicatrization or any use where a hemostatic and/or bacteriostatic and/or aesthetic effect is sought. In fact, fibrinogen, a major constituent in such a concentrate, undergoes enzymatic degradation in contact with thrombin activated by calcium ions. After elimination of the fibrinopeptides A and B, the fibrin monomers polymerize and spontaneously generate soluble fibrin. The presence of Factor XIII in this type of product contributes to stabilizing the fibrin by covalent bonds by making it insoluble, i.e. resistant to solvent (e.g., urea). The fibrin thus stabilized, in addition to its coagulant role, is more resistant to fibrinolysis and to mechanical tractia.
For these different therapeutic uses, it is necessary to have available a product of plasmatic origin with good solubility and stable under the conditions of use. In so far as biological glues are concerned, it is further necessary for them to have, after being placed in contact with the calcic thrombin, a high adhesive power and high elasticity.
The fact of obtaining such characteristics is directly related to the nature of the product and so to the method of purification thereof from plasma. It is then useful to have a preparation technique readily usable on an industrial scale but which is also sufficiently gentle so as not to impair the biochemical qualities of the product for the desired use by clinicians.
Glues containing fibrinogen and Factor XIII are already known particularly from French patents No. 2,448,900 and 2,448,901. These products are obtained from plasma cryoprecipitate by treating with a buffer solution containing plasminogen inhibitor-activator or plasmin inhibitor, which is present in the glue in the lyophilized state.
These products have interesting characteristics. However, these products are obtained using fairly complex fibrinogen production methods which require, so as to have a final satisfactory mixture, the exogenous addition of other plasmatic proteins such as Factor XIII. Moreover, during production, inhibitors must be added such as protease inhibitors of animal origin such as bovine aprotinin.
Furthermore, the known protein concentrates, in particular those used as glues, do not solubilize in an aqueous medium at ambient temperature and may even be difficult to solubilize at 37.degree. C., making it necessary to add to the lyophilization vial of the product a magnetized bar and provide a magnetic agitator used for accelerating the solubilization.
It would then be quite beneficial to have available a simple method of producing biological glue so as to obtain without exogenous addition of proteins, a protein mixture corresponding to the desired qualities for such a product. In particular, for the use as glue, the concentrate obtained should have satisfactory fibrinogen, Factor XIII and fibronectin content.
The method in question must also be readily adaptable to modifications for introducing a viral inactivation step. The resultant product should solubilize in less than 10 minutes at the temperature of use (generally 18.degree. to 20.degree. C.) without requiring special tools. The product should also be stable for several hours so as to facilitate use and guarantee the clinical efficiency thereof.